The cell responsible for bone formation is the osteoblast. Recent work indicates that osteoblasts are deleted from bone by an evolutionarily conserved physiological process called apoptosis. Activation of the apoptotic program is critical for the maintenance of tissue size, shape and function; interference with apoptosis is tightly linked to craniofacial, limb and spine malformations. Surprisingly, little is known of the regulation of the apoptotic program in bone. The overall hypothesis of the present grant proposal is that a local change in pO2 and/or Pi can trigger osteoblastic apoptosis, by altering the cell's oxidative metabolism (resulting in a loss of membrane potential and the induction of 'permeability transition' state), activating Bcl-2 and enhancing caspase activity. These changes, the principal investigator believes, will alter the expression of redox-sensitive transcription factors, NF-kB and AP-1. To assess this postulation, four Specific Aims are outlined. In Specific Aim 1, the applicant proposes to determine how Pi and pO2 changes regulate osteoblast apoptosis in cell cultures and calvaria. In Specific Aim 2, he proposes to ascertain whether the induced apoptosis is mediated by alterations in the expression and/or activity of members of the Bcl superfamily, specifically with respect to homodimer and heterodimer formation between Bcl-2, Bcl-XL and Bcl-XS, as well as changes in Bcl-2 localization. In Specific Aim 3, the principal investigator seeks to determine if changes in mitochondrial function promote osteoblastic apoptosis and an induction in the activity of caspases, a family of cysteinyl aspartate proteinases that are acknowledged to have a role in apoptosis. Finally, in Specific Aim 4, the applicant intends to study the effect of Pi and a low PO2 on the expression and function of two redox-sensitive transcription factors, namely, NF-kB and AP-1. Taken together, these studies should highlight the molecular basis of bone cell apoptosis, a phenomenon that is now considered critical for bone growth and skeletal remodeling.